Neisseria gonorrhoeae has a single reservoir--Homo sapiens; mankind, in turn, possesses multiple mechanisms with which to fight infection. Despite this, untreated gonorrhea can persist for weeks. We have shown that Por protein of N. gonorrhoeae (Por) inhibits granule enzyme release (LER), but not O2 generation after chemoattractants (CTX). Por has no effect on LER triggered by immune complexes (IC). We have proposed that Por prevents phagosome-lysosome fusion in neutrophils (PMN) prolonging gonococcal survival early in the course of infection. We will examine this as follows: As Por- mutants are unavailable, we will determine the effect of Por added to bactericidal assays of E. Coli transfected with opa, the gonococcal opacity-associated and attachment protein. We will determine whether Por inhibits LER, O2 release or phagocytosis triggered by opa +E. coli or opa+ liposomes. We have demonstrated that Por inhibits production of diacylglycerol (DAG) and metabolism of phosphatidylcholine (PC) in PMN treated with CTX but does not affect generation of phosphatidate (PA). LER is tightly correlated with production of DAG, most likely via a PC-specific phospholipase C (PC-PLC) which is inhibited by Por. In parallel to its functional effects, Por does not inhibit DAG generation triggered by IC. We will then focus on the role of Por and opa of the gonococcus in affecting signal transduction in PMN and how this may favor neisserial infection. We will determine where Por is located after it binds to PMN (plasmalemma cytoskeleton, granules) using light and electronmicroscopic techniques (fluorescent antibody immunogold-labelling-125I-labelling and cell fractionation). We will determine whether this inhibition is secondary to porin activity by determining whether, in fact, Por acts as an ion channel in vitro (ion fluxes [22Na, 36Cl-, 42K+ ,35SO-2-4] or fluorescent probes (SBFI, PBFI, or SPQ). We will determine whether CTX provoke metabolism of both the diacyl- and alkyl,acyl (EA) fractions of phosphatidylcholine (radiolabel, phospholipid mass after CTX) and whether EAG generation is inhibited by Por. We will perform similar experiments in PMN triggered by opa+ E. coli and opa+ liposomes. We will determine whether PC-PLC activity in purified plasma membranes can be augmented by treatment with the above CTX. We will attempt to parallel the effects of Por by using other disruptors of ion movements (amiloride, SITS, DIDS) and determine PL turnover (HPLC,TLC). We will determine whether other organelles are required for activation and inhibition of PC-PLC by examining PL metabolism in cytoplasts. We will determine the effect of Por on phospholipid domains of the cell membrane (freeze fracture). We will examine whether activation of the PC-PLC universally triggers exocytosis after CTX in PMN and monocytes. We will determine the source of DG after IC by examining diradyl-PL metabolism. In learning how the gonococcus can escape immediate killing, we will learn both how to treat this disease and how to control pathologic inflammation.